Investigating the transcriptional control of cardiovascular development

Abstract

Transcriptional regulation of thousands of genes instructs complex morphogenetic and molecular events for heart development. Cardiac transcription factors choreograph gene expression at each stage of differentiation by interacting with cofactors, including chromatin-modifying enzymes, and by binding to a constellation of regulatory DNA elements. Here, we present salient examples relevant to cardiovascular development and heart disease, and review techniques that can sharpen our understanding of cardiovascular biology. We discuss the interplay between cardiac transcription factors, cis-regulatory elements, and chromatin as dynamic regulatory networks, to orchestrate sequential deployment of the cardiac gene expression program.

Keywords: cell differentiation; heart diseases; transcription factors; transcriptional networks; transcriptional regulatory elements.

Figure 1. Molecular players for transcriptional regulation

(A) Cis-regulatory elements containing DNA binding sites are bound by transcription factors and (B) modulate the assembly of the Pre-Initiation Complex at promoters through (C) physical contacts driven by a three-dimensional arrangement of chromatin, thereby acting as (D) a molecular platform between cellular signaling and gene activity.

Figure 2. Studying cardiac regulatory elements

(A) (Left) Evolutionary constraints on some regulatory elements render them identifiable by comparative genomics, as exemplified by enhancers upstream of mouse Nkx2-5 . (Right) Combinations of specific chromatin features can reveal potential regulatory elements active in a given cell type. (B) (Left) Enhancer activity is classically tested by an ability of candidate elements to drive tissue- or stage-specific activity of a reporter. (Middle) New technologies, such as SIF-seq , allow screening of large genomic neighborhoods for tissue-specific enhancers. (Right) GROMIT is a technology that can reveal integrated regulatory inputs exerted at a locus.

Figure 3. Developmental patterning of regulatory elements

(A, B) Dynamics of transcriptional activity can result from stage-specific action of transactivators or repressors. (C) Cellular history of the genome can also influence transcriptional responses, such as by epigenetic priming of regulatory elements.

Figure 4. Building regulatory circuits to implement developmental patterning

Simple network motifs allow the emergence of temporal patterning for transcriptional responses. (A) Positive feedback, direct or indirect, can establish cellular memory of an exposure from transient events. (B) An incoherent feed-forward loop can generate a transient expression pulse. (C) TFs and cis-regulatory elements are molecular building blocks of transcriptional networks. As TFs control activity of other TF genes, they create functional connections that organize into regulatory modules, which can be assembled into complex stage- and tissue-specific regulatory networks for transcriptional patterning.

Figure 5. Population vs. single-cell approaches for gene expression

Insights from cell population–based studies reveal global changes in fields of cells or tissue during cardiac differentiation and development. However, cell heterogeneity can obscure gene expression analysis from cell populations. Two distinct modes of transcriptional responses, rheostatic (or analog) and binary (or digital), give similar mean behaviors on a population scale. Yet, in a rheostatic regime, cells respond in a concerted fashion homogeneously, while a binary regime leads to the appearance of populations with expressers or non-expressers. During development, this can have fundamental consequences on how cells differentiate. Of importance, long-range activation by enhancers acts through a binary mechanism, therefore contributing of cell-to-cell heterogeneity .